In the United States there are at least eleven serious diseases that are transmitted by ticks. Current diagnostic testing methods for these disease causing pathogens are limited and/or unreliable. Furthermore, no test exists that can cover the breadth of pathogens transmitted by ticks. In our Phase I proposal, we plan to develop a tick-borne pathogen detection assay for use on the Ibis T5000 system. This system employs broad-range PCR and electrospray ionization mass spectroscopy (PCR/ESI-MS). The assay to be developed and validated in this proposal will identify and speciate broad groups of bacteria, including all groups that contain known tick- borne bacteria and flaviviruses. As our assay is broad range, any novel tick-borne bacterial/viral pathogens present will also likely be detected. To achieve our goals, we will employ three key technologies: 1) Broad- range PCR primers and ESI-MS to detect a wide variety of tick-borne pathogens that might be missed by specific PCR 2) A mechanical cell destruction method, shown to lyse even the most durable forms of bacteria, such as Bacillus anthracis spores, which may enable the identification of bacteria that previously have not been detected and increase the amount of detectable pathogen DNA 3) Whole genome amplification to increase trace levels of pathogen DNA to levels detectable by PCR. In Phase I the assay will be validated on a large number of field-collected ticks from Lyme endemic areas. In Phase II, we plan to evaluate the effectiveness of our assay on human clinical samples (e.g., erythema migrans skin biopsies, cerebral spinal fluid, synovial fluid, whole blood). Upon the completion of Phases I and II, Ibis Biosciences will make available to the healthcare community a validated assay to detect bacteria/viruses in ticks and human clinical samples. Results of this work will lead to a commercially available diagnostic to detect and identify tick-borne diseases in humans for improved medical treatment and public health surveillance. [unreadable] [unreadable] In the United States there are at least eleven serious diseases that are transmitted by ticks. Current diagnostic testing methods for these disease causing pathogens are limited and/or unreliable. Furthermore, no test exists that can cover the breadth of pathogens transmitted by ticks. In this Phase I SBIR proposal we will develop and validate a broad-range tick-borne pathogen detection assay using field collected ticks. In Phase II we will continue this work using human clinical samples. Results of this work will lead to a commercially available diagnostic to detect and identify tick-borne diseases in humans for improved medical treatment and public health surveillance. [unreadable] [unreadable] [unreadable]